The long-term goal of this project is to understand how human genetic variation modifies drug-drug interactions. We focus here on metabolic interactions involving the widely used oral anticoagulant drug, warfarin. Warfarin has, for many decades, provided a model system for studying mechanisms of drug-drug interactions, but no information is available as to the extent to which common polymorphisms in warfarinresponse genes affect these drug interactions. CYP2C9 genotype is particularly important because this enzyme governs the metabolic inactivation of the more potent S-enantiomer of the drug. Consequently, the current proposal addresses the central question;How does CYP2C9 genotype modify warfarin drug interactions that have a metabolic basis? This will be accomplished with the following specific aims; Aim 1 Determine the magnitude of the warfarin-fluconazole inhibitory drug interaction in healthy volunteers genotyped for CYP2C9*1/ *1, *1/*3 or *3/*3 by measuring warfarin enantiomer AUC, clearance, partial metabolite clearances, and prothrombin time. Aim 2 Determine the magnitude of the warfarin-rifampin inductive drug interaction in the same genotyped subject population studied in Aim 1. Aim 3 Exploit CYP2C9 null systems to determine the role of alternative P450s in S-warfarin clearance in subjects with genetically compromised CYP2C9 activity, and develop in vitro systems to recapitulate the in vivo metabolic interactions characterized in Aims 1 and 2. Aim 4 Synthesize known and suspected inhibitory metabolites of amiodarone, and determine their inhibitory potency in vitro against P450 enzymes known to contribute to warfarin clearance. Aim 5 Define, in warfarin patients, the contribution of CYP2C9 genotype, CYP2C8 genotype and steadystate plasma levels of amiodarone and its inhibitory metabolites, to variability in warfarin dose adjustment in patients receiving combination therapy. Aims 1 and 2 address the hypothesis that functionally defective CYP2C9 alleles attenuate the warfarinfluconazole inhibitory interaction and exacerbate the warfarin-rifampin inductive interaction. Aim 3 addresses the hypothesis that CYP2C9-deficient subjects are at increased risk from metabolic interactions involving CYP3A4. Aims 4 and 5 address the hypothesis that the magnitude of the warfarin-amiodarone drug interaction can be predicted using information about the genotype status for CYP2C9, CYP2C8, and the plasma concentration of circulating inhibitory metabolites of amiodarone. Successful completion of these studies is expected to improve clinical care of warfarin patients undergoing treatment with P450 inhibitors and inducers by providing a mechanistic framework, incorporating pharmacogenomic considerations, that will improve guidance of dose adjustment during polytherapy with this widely used oral anticoagulant.